Purpose:
Age-related macular degeneration (AMD) is a progressive neurodegenerative disease that is the leading cause of blindness in elderly individuals in developed countries. Risk for AMD is mediated by genetic and environmental factors; known major risk loci include ARMS2/HTRA, CFH, C2/CFB, and C3. These loci harbor common genetic variants that influence risk for AMD; rare variation has not been extensively examined in these regions. We hypothesized that rare variants contribute to AMD susceptibility.

Methods:
We evaluated individuals from the Amish communities of Ohio and Indiana, an isolated population useful for genetic studies due to the relatively few founders, limited heterogeneous environmental exposure, and the presence of detailed records on their multigenerational pedigrees. Genetic information from this population can be used to further the knowledge of diseases know to have genetic and environmental components, including AMD. Exome sequencing was performed in five members of a small nuclear Amish family with AMD who lack the common risk alleles at the major AMD risk loci. Follow-up genotyping and association analysis using MQLS was performed in a cohort of 929 additional Amish individuals including 95 with self-reported AMD (15 confirmed); self-report of AMD is a reliable proxy for AMD disease status in this cohort.

Results:
Exome sequencing identified a variant (P503A) in CFH that is not present in dbSNP or 1000Genomes and is predicted to be damaging by Polyphen2. Further, the variant has a GERP score of 3.64, indicating the base pair is strongly conserved across species. Three additional rare variants were detected in MFN2, RIC8B, and LRP4, genes which are not known contributors to AMD, but which warrant further follow-up to determine potential undetected involvement. Analysis of 900 additional members of an Amish cohort revealed that the four variants were associated (P<0.005) with AMD, with P503A being the most significantly associated (P<1x10-6). P503A was absent when evaluated in a cohort of 700 elderly non-AMD, non-Amish controls.

Conclusions:
Exome sequencing of five members of an Amish family lacking the common AMD risk alleles identified four novel variants that appear to contribute to AMD. Identification of these novel risk variants provides evidence that rare variants remain to be detected in AMD and that these variants can be identified through exome sequencing of major AMD risk loci.